Propylene is an important basic raw material of synthetic resins such as polypropylene and petrochemical products and used in a wide variety of products such as automobile bumpers, food containers, films, and medical instruments.
Isopropyl alcohol produced from a plant-derived raw material can be converted to propylene through a dehydration process. Therefore, isopropyl alcohol is promising as a raw material for carbon-neutral propylene. At present, Kyoto Protocol mandates that developed countries as a whole reduce carbon dioxide emissions by 5% as compared to 1990 levels in the 2008 to 2012 period. Therefore, carbon-neutral propylene is extremely important in the global environment because of its versatility.
Bacteria that assimilate a plant-derived raw material to produce isopropyl alcohol are already known. For example, the pamphlet of WO2009/008377 discloses a bacterium modified to produce isopropyl alcohol from glucose as a raw material and described that the bacterium has excellent properties as a biocatalyst for industrial production because of its high selectivity of isopropyl alcohol.
In an isopropyl alcohol-producing Escherichia coli, since the raw material for isopropyl alcohol is glucose, a large number of compounds obtained by glycolysis and catabolism can all be by-products. On the other hand, since those compounds may be substances essential for growth of Escherichia coli, it is impossible to completely suppress the amount of glucose consumed by those secondary reactions. Accordingly, to increase the production rate of isopropyl alcohol while minimizing by-products, it is necessary to maximize metabolic flow to isopropyl alcohol while considering all metabolic reactions occurring in Escherichia coli, and various techniques have been proposed from the viewpoint of biological activity and substance production.
For example, the pamphlet of WO2009/008377 discloses an isopropyl alcohol-producing bacterium in which respective genes of acetoacetate decarboxylase, isopropyl alcohol dehydrogenase, CoA transferase, and thiolase are introduced to allow isopropyl alcohol to be produced from a plant-derived raw material. It is described that the isopropyl alcohol-producing bacterium can achieve a production rate of 0.6 g/L/hr and an amount of accumulation of 28.4 g/L.
The pamphlet of WO2009/049274 and Appl. Environ. Biotechnol., 73(24), pp. 7814-7818, (2007) disclose Escherichia coli in which respective genes of acetyl-CoA acetyltransferase, acetoacetyl CoA transferase, acetoacetate decarboxylase, and secondary alcohol dehydrogenase are introduced to produce isopropyl alcohol. It is described that those bacteria can achieve a production rate of 0.4 g/L/hr, a yield of 43.5%, and an amount of accumulation of 4.9 g/L.
The pamphlet of WO2009/028582 discloses Escherichia coli in which respective genes of acetoacetate carboxylase, isopropyl alcohol dehydrogenase, acetyl-CoA:acetate CoA-transferase, and acetyl-CoA acetyl transferase are introduced to produce isopropyl alcohol. It is described that the bacterium can achieve an amount of accumulation of 9.7 g/L.
Appl. Microbial. Biotechnol., 77(6), pp. 1219-1224, (2008) discloses Escherichia coli in which respective genes of thiolase, CoA-transferase, acetoacetate decarboxylase, and primary-secondary alcohol dehydrogenase are introduced to produce isopropyl alcohol. It is described that the bacterium can achieve a production rate of 0.6 g/L/hr, a yield of 51%, and an amount of accumulation of 13.6 g/L.
The pamphlet of WO2009/103026 discloses Escherichia coli in which respective genes of acetoacetate decarboxylase, acetyl-CoA:acetate CoA transferase, acetyl-CoA acetyl transferase, and isopropyl alcohol dehydrogenase are introduced to allow the production of isopropyl alcohol. It is described that the bacterium is expected to have the ability to achieve a yield of 50%, a production rate of 0.4 g/L/hr, and an ultimate production of 14 g/L.
The pamphlet of WO2009/247217 discloses Escherichia coli in which respective genes of acetoacetate decarboxylase, CoA transferase, thiolase, and 2-propyl alcohol dehydrogenase are introduced to allow the production of isopropyl alcohol. It is described that the bacterium can achieve an ultimate production of 2 g/L.
Here, isopropyl alcohol dehydrogenase, secondary alcohol dehydrogenase, primary-secondary alcohol dehydrogenase, and 2-propyl alcohol dehydrogenase are enzymes that have different names but catalyze the same reaction. CoA transferase, acetoacetyl CoA transferase, acetyl CoA:acetate CoA transferase, and CoA-transferase are enzymes that have different names but catalyze the same reaction. In addition, acetoacetate decarboxylase and acetoacetate decarboxylase are enzymes that have different names but catalyze the same reaction, and thiolase and acetyl CoA acetyl transferase are enzymes that have different names but catalyze the same reaction. Accordingly, although the productivity of the isopropyl alcohol-producing Escherichia coli of the above-described documents varies, the enzymes used to produce isopropyl alcohol are equivalent to the four enzymes—acetoacetate decarboxylase, isopropyl alcohol dehydrogenase, CoA transferase, and thiolase—described in the pamphlet of WO2009/008377. For purposes such as improvement of productivity and yield, the four enzymes have been conventionally studied.
On the other hand, a method of deleting an enzyme malate dehydrogenase that a microorganism possesses is known as a method for improving the yield and productivity in substance production by the microorganism.
For example, the pamphlet of WO2009/023493 describes that, in the production of 1,4-butanediol by Escherichia coli, the yield is increased by disruption of a malate dehydrogenase gene that the Escherichia coli possesses or by simultaneous disruption of a malate dehydrogenase gene and a transhydrogenase gene that the Escherichia coli possesses.
Additionally, the pamphlet of WO2009/012210 describes that, in the production of ethanol by Escherichia coli, the yield is increased by simultaneously disrupting a malate dehydrogenase gene and a D-lactate dehydrogenase gene that the Escherichia coli possesses.
Furthermore, the pamphlet of WO2009/111672 describes that, in the production of dodecanol by yeast, productivity is effectively improved by simultaneously disrupting acetaldehyde-CoA dehydrogenase gene, a D-lactate dehydrogenase gene, and a malate dehydrogenase gene that the yeast possesses.